Optical fibers guide light between separate locations and enable new types of fluorescence imaging. Fiber-optic fluorescence imaging systems include portable handheld microscopes, flexible endoscopes well suited for imaging within hollow tissue cavities and microendoscopes that allow minimally invasive high-resolution imaging deep within tissue. A challenge in the creation of such devices is the design and integration of miniaturized optical and mechanical components. Until recently, fiber-based fluorescence imaging was mainly limited to epifluorescence and scanning confocal modalities. Two new classes of photonic crystal fiber facilitate ultrashort pulse delivery for fiber-optic two-photon fluorescence imaging. An upcoming generation of fluorescence imaging devices will be based on microfabricated device components.Fiber-optic fluorescence imaging has become increasingly versatile over the last decade as fiber-based devices have declined in size but gained in functionality. Three classes of applications in live animal and human subjects provide the primary motivations for innovation in fiber-optic imaging. First, basic research on biological and disease processes would benefit enormously from instrumentation that permits cellular imaging under conditions in which conventional light microscopy cannot be used. Many cell types reside within hollow tissue tracts or deep within solid organs that are inaccessible to optical imaging without devices that can reach such locations in a minimally invasive manner. Flexible fiber-optic devices also allow handheld imaging and imaging in freely moving animals 1 . Second, fiber devices might be implanted within live subjects for long-term imaging studies. This capability will not only benefit research on the cellular effects of aging, development or experience, but also will lead to new in vivo assays for testing of drugs and therapeutics. By allowing examination of cells concurrently with observation of disease symptoms and animal behavior, fiber-optic imaging will permit studies correlating cellular properties and disease outcome in individual subjects over time and could reduce the numbers of animals needed. A third set of applications concerns development of minimally invasive clinical diagnostics and surgical procedures 2 . Although much work remains, fiber-optic instrumentation has already broadened the applicability of in vivo fluorescence imaging.The seeds for development of fiber optic in vivo imaging were planted by early uses of fiberoptics and fluorescence for chemical sensing and spectroscopy, including classic work on detection of intracellular redox states [3][4][5] . Use of fiber optics for remote sensing and spectroscopy remains strong today 6 , and has expanded to include bioluminescence detection 7 . The recent proliferation of fluorescent probes has widened the set of potential uses © 2005 Nature Publishing Group Correspondence should be addressed to M.J.S (mschnitz@stanford.edu). COMPETING INTERESTS STATEMENTThe authors declare that they have no co...

One of the major limitations in the current set of techniques available to neuroscientists is a dearth of methods for imaging individual cells deep within the brains of live animals. To overcome this limitation, we developed two forms of minimally invasive fluorescence microendoscopy and tested their abilities to image cells in vivo. Both one- and two-photon fluorescence microendoscopy are based on compound gradient refractive index (GRIN) lenses that are 350-1,000 microm in diameter and provide micron-scale resolution. One-photon microendoscopy allows full-frame images to be viewed by eye or with a camera, and is well suited to fast frame-rate imaging. Two-photon microendoscopy is a laser-scanning modality that provides optical sectioning deep within tissue. Using in vivo microendoscopy we acquired video-rate movies of thalamic and CA1 hippocampal red blood cell dynamics and still-frame images of CA1 neurons and dendrites in anesthetized rats and mice. Microendoscopy will help meet the growing demand for in vivo cellular imaging created by the rapid emergence of new synthetic and genetically encoded fluorophores that can be used to label specific brain areas or cell classes.

A central goal in biomedicine is to explain organismic behavior in terms of causal cellular processes. However, concurrent observation of mammalian behavior and underlying cellular dynamics has been a longstanding challenge. We describe a miniaturized (1.1 g mass) epifluorescence microscope for cellular-level brain imaging in freely moving mice, and its application to imaging microcirculation and neuronal Ca2+ dynamics.

SUMMARY A major technological goal in neuroscience is to enable the interrogation of individual cells across the live brain. By creating a curved glass replacement to the dorsal cranium and surgical methods for its installation, we developed a chronic mouse preparation providing optical access to an estimated 800,000–1,100,000 individual neurons across the dorsal surface of neocortex. Post-surgical histological studies revealed comparable glial activation as in control mice. In behaving mice expressing a Ca2+ indicator in cortical pyramidal neurons, we performed Ca2+ imaging across neocortex using an epi-fluorescence macroscope and estimated that 25,000–50,000 individual neurons were accessible per mouse across multiple focal planes. Two-photon microscopy revealed dendritic morphologies throughout neocortex, allowed time-lapse imaging of individual cells, and yielded estimates of >1 million accessible neurons per mouse by serial tiling. This approach supports a variety of optical techniques and enables studies of cells across >30 neocortical areas in behaving mice.